Growth of plants is greatly influenced by moisture conditions of soil. In order to adapt themselves to conditions where moisture in soil is lacking and to maintain their growth, plants adopt a strategy of drought resistance via drought avoidance or physiological tolerance. Drought avoidance refers to a property, for example, of avoiding drought stress by deeply elongating their roots into the soil or resting during periods when moisture is lacking in the soil. Physiological tolerance to drought refers to a property of plants which allows them to grow even in dry environments.
Currently, farms are located mostly in semiarid zones around the world, and crop production is greatly limited by the amount of water available. In soil, the moisture content is often relatively high at a greater depth as compared to soil closer to the surface because drying of soil starts from the surface and proceeds downwards. Thus, drought avoidance mediated by a trait which allows plants to more deeply elongate their roots into the ground, i.e., deep root growth (deep rooting property), among other adaptation strategies of plants against drought conditions is expected to greatly contribute to growth maintenance and increased yield of plants in drought regions. Drought avoidance by plants is also helpful in conservation of the environment globally. To develop deep-rooted plants, a large number of plants must be evaluated, but it is difficult to evaluate roots of a large number of individual plants by direct observation because roots exist in the soil, therefore there is a need for developing more convenient and efficient methods for evaluating deep root growth (deep rooting property) of plants.
Several methods for examining plants for deep root growth (deep rooting property) have been known, including the trench method, cylinder method, etc.
According to the trench method, plants are cultivated in a field and a trench of about 0.5 m to 2 m in depth is dug to examine the thickness and number of roots of the plants at different depths in the trench (Nemoto, H., Suga, R., et al., Breeding Science 48: 321-324, 1998). The trench method requires much labor for digging a trench and involves difficulty in evaluating a large number plants. Moreover, soil moisture conditions are influenced by natural weather because plants are cultivated in a field. Individuals, lines or varieties evaluated as rapidly elongating their roots under sufficient soil moisture conditions do not always rapidly elongate their roots under drought conditions, because it may be highly possible that the root growth speed varies with soil moisture conditions, i.e., moisture content. Thus, even if such labor-intensive examination is used, there exists a risk of selecting varieties/lines which are poor at elongating their roots under drought conditions.
According to the cylinder method, a plant is cultivated in a cylinder made from plastic or the like and the growth of roots is examined.
An example of the cylinder method comprises cultivating a plant in a transparent plastic cylinder inclined to allow roots to appear on the lateral face and measuring the growth speed or maximum depth of the roots (Mia, M. W., Yamauchi, A., et al., Japanese Journal of Crop Science, 13: 131-140, 1996). This method has the advantage that roots can be directly observed, but only one plant can be examined in a cylinder because it cannot be known for certain which root originates from which plant if multiple plants are planted in one cylinder.
Trillana et al. sowed rice seeds in a pot of 1 m in length made from polyvinyl chloride, followed by submerged cultivation at a water level of 2 or 3 cm above the surface of soil for 14 days after sowing, and partially drained water on 62 days after sowing to lower the water level to 30 cm from the bottom of the pot (60 cm below the surface of soil), and continued cultivation for 6 days, followed by submerged cultivation at a water level of 2 or 3 cm above the surface of soil again, and compared the densities of roots and dry weights of leaves and stems (Trillana, N., Inamura, T., et al., Plant Production Science 4: 155-159, 2001). The method of Trillana et al. is not suitable for large-scale screening because it is thought that much labor is required to adjust the water level by replenishing each vessel with water everyday to keep the water level constant after drought stress treatment. Moreover, the method of Trillana et al. requires 4 hours to drain water for drought stress treatment. The water level is lowered only once in this test, which is very inconvenient when the water level is to be gradually lowered in several steps. If water is added from above to fill a tall vessel packed with soil, it is difficult to remove the air in the soil and a considerable time is required until water penetrates into the soil. Thus, the method of Trillana et al. is suitable for basic studies dealing with few tested plants, but unsuitable for breeding, which requires analyses of a large number of individuals.
In both cylinder methods disclosed by Mia et al. and Trillana et al., different varieties are cultivated in separate vessels, and therefore, the growth of roots of multiple varieties is not compared under the same soil moisture conditions. The soil moisture content in a culture vessel always varies with the amount of water absorbed by the plant. The amount of water absorbed by a plant varies with environmental conditions and growth stage as well as the size of the plant and the like, so that the soil moisture content is greatly influenced by these factors. If different varieties are planted in separate vessels, they cannot be compared under the same soil moisture conditions because varieties of larger plants undergo stronger drought stress earlier than varieties of smaller plants even under the same temperature or light conditions. In fact, the report of Trillana et al. shows that the soil moisture content at the end of drought stress treatment varied between the varieties cultivated, allegedly because of the difference in the growth of aerial parts. As described above, they kept the water level constant after drought stress treatment by replenishing each vessel with water everyday, but this operation requires much labor and if this water level adjustment were omitted, it would be certain that the soil moisture content would vary more widely between vessels, and this difference appears to greatly influence test results. Thus, deep rooting property cannot be compared under the same soil moisture conditions when multiple varieties or lines are cultivated in separate vessels because the soil moisture conditions vary between the vessels.
A method for evaluating drought resistance of plants rather than deep rooting property using plastic pots has been proposed (JPA 2003-230318). This method comprises growing a test plant in multiple pots under suitable cultivation conditions for a short period, then stopping irrigation to perform drought stress treatment, followed by cultivation under sufficient irrigation for a given period by varying the irrigation period pot by pot, and then examining the survival rate in relation to a period of drought stress treatment. In this method for evaluating drought resistance it is also shown that the soil moisture content varies between pots over time after drought stress treatment because only one type (variety) of plant is grown in one pot. Thus, the survival rate widely differs between two varieties having the same drought resistance limit even if they are treated drought stress for the same number of days. This method is very complicated because it involves preparing multiple pots for each tested plant and measuring the soil moisture content for each period to compare the survival rate at the same soil moisture content. Thus, the drought resistance test using pots also causes a difference in soil moisture content when different plants are cultivated in separate vessels, which makes it difficult to compare drought resistance between the plants.
A method for evaluating drought resistance in plants by using relatively shallow pots has also been reported (Wada, Suzuki, et al., Japanese Journal of Crop Science, 70: 580-587, 2001). This method comprising growing multiple test plants in one pot under suitable cultivation conditions for a short period, then stopping irrigation to perform drought stress treatment, and cultivating the plants under sufficient irrigation again for a certain period, wherein the growth state of each plant is examined at various instants. This method does not require much labor and allows multiple plants to be compared at the same soil moisture content, but it “compares drought resistance under low soil moisture conditions” and has little relevance to the trait allowing plants to elongate their roots deeply into soil to avoid drought (i.e., deep rooting property) (Nemoto, H., Suga, R., et al., (1998) Breeding Science 48; 321-324) and plants selected by this method are not always deep-rooted.
In order to develop deep-rooted plants, it is necessary to test a number of individuals or lines, but conventional methods for evaluating plants for deep rooting property as described above are unsuitable for testing a large number of tested plants under the same soil moisture conditions. There is a need for methods for efficiently and conveniently evaluating/screening a large number of plants for deep rooting property under the same soil moisture conditions wherein cultivation conditions can be readily controlled, e.g., the soil moisture content can be readily adjusted.    Patent document 1: JP 2003-230318 A    Non-patent document 1: Nemoto, H., Suga, R., et al., Breeding Science 48: 321-324, 1998    Non-patent document 2: Mia, M. W., Yamauchi, A., et al., Japanese Journal of Crop Science, 13: 131-140, 1996    Non-patent document 3: Trillana, N., Inamura, T., et al., Plant Production Science 4: 155-159, 2001    Non-patent document 4: Wada, Suzuki, et al., Japanese Journal of Crop Science, 70: 580-587, 2001.